A heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a protein belonging to the epidermal growth factor (EGF) family. It has been revealed that this protein promotes cell proliferation, differentiation, chemotaxis, and so forth by binding to an EGF receptor (EGFR/ErbB1 or ErbB4) (NPLs 1 to 3).
Moreover, it has been revealed that HB-EGF in vivo contributes to myogenesis, heart development, and wound healing. This suggests that HB-EGF is an important factor in organogenesis (NPLs 4 to 6).
Further, there have been reports that HB-EGF is involved in cancer proliferation and progression at a variety of aspects such as the proliferation of pancreatic cancer (NPL 7), the proliferation of stomach cancer (NPL 8), the association with skin cancer (NPL 9), the association with drug resistance in head and neck cancers (NPL 10), and angiogenesis to cancer tissues (NPL 11). It has been revealed that HB-EGF is an important factor also in various cancers.
Moreover, it has been revealed that HB-EGF is first synthesized in the form of a type I transmembrane protein (transmembrane HB-EGF), and then an extracellular region immediately above a cell transmembrane portion in the cell membrane is cleaved by a protease and subsequently released in the form of 14- to 22-kilodalton soluble HB-EGF (NPLs 12 and 13). Further, it is also known that soluble HB-EGF formed by such cleavage functions as a growth factor which activates EGFR/ErbB1 of its own HB-EGF expressing cell in an autocrine manner, or which activates EGFR/ErbB1 of the other cells in a paracrine manner.
On the other hand, it is known that transmembrane HB-EGF itself also functions as a growth factor which activates EGFR/ErbB1 of other nearby cells in a juxtacrine manner (NPL 14). However, it has been shown that the cell proliferating activity of transmembrane HB-EGF is weaker than that of soluble HB-EGF (NPL 11). These results suggest the process of forming soluble HB-EGF by protease cleavage is important in making HB-EGF exhibit the function as a growth factor.
Meanwhile, mutant mice prepared to express transmembrane HB-EGF but not to express soluble HB-EGF have been produced by introducing an amino acid mutation into a cleavage site of HB-EGF. Then, in the analysis of the mutant mice, abnormal heart organogenesis has been observed as in the case of knockout mice which express no HB-EGF at all (NPLs 5 and 15). Further, it has also been shown that suppressing the HB-EGF cleavage by a protease inhibitor suppresses cardiac hypertrophy due to soluble HB-EGF (NPL 16). This suggests that soluble HB-EGF plays a role in the aforementioned important physiological function of HB-EGF in the organogenesis.
Moreover, it is also known that, in a cancer, a cleaved intracellular region (HB-EGF-CTF) of HB-EGF is translocated to the nucleus and functions to promote the cell division (NPL 17). Further, it has been shown that suppressing the HB-EGF cleavage by a protease inhibitor can inhibit the proliferation and invasion of stomach cancer (NPL 18). Hence, it has been revealed that the HB-EGF cleaving process is an important factor in the proliferation and so forth of cancer cells, too.
Based on the above-described findings, researches and developments have been in progress for treatment methods against various diseases (such as heart failure, nerve diseases, lung diseases), particularly cancers, in which HB-EGF is presumably involved. For example, the anti-tumor effect of an anti-HB-EGF antibody or the like has been confirmed in a xenograft mouse model obtained by transplanting ovarian cancer cells (NPLs 19 and 20). Further, it has also been shown that the proliferation of cancer cells can be inhibited by inhibiting the HB-EGF cleavage using an antibody to suppress soluble HB-EGF formation (NPL 21). As such, antibodies have been developed which are capable of binding to HB-EGF and exhibiting such activities as anti-tumor activity and cleavage inhibitory activity.
As described above, in order for an antibody against HB-EGF to have sufficient activities in the treatments against cancers or the like, in other words, to completely block signal transduction in cell proliferation or the like in which HB-EGF is involved, it is presumably necessary that such an antibody should have both of two activities: (1) strongly inhibiting the cleavage that would lead to the formation of soluble HB-EGF and HB-EGF-CTF functioning as a growth factor or the like (cleavage inhibitory activity); and (2) inhibit binding of soluble HB-EGF and transmembrane HB-EGF to an EGF receptor (EGFR/ErbB1 or ErbB4), consequently strongly inhibiting the activation and so forth of the EGF receptor (neutralizing activity).
Nevertheless, for example, the antibody described in NPL 21 has a cleavage inhibitory activity as described above, but this literature has also revealed that the antibody does not have a neutralizing activity. Judging from the foregoing, under current situations, no antibody against HB-EGF has been developed which has sufficient activities in the treatments of diseases, particularly cancers, in which HB-EGF is involved.